Growth Hormone Receptor Regulation in Cancer and Chronic Diseases

The GHR signaling pathway plays important roles in growth, metabolism, cell cycle control, immunity, homeostatic processes, and chemoresistance via both the JAK/STAT and the SRC pathways. Dysregulation of GHR signaling is associated with various diseases and chronic conditions such as acromegaly, cancer, aging, metabolic disease, fibroses, inflammation and autoimmunity. Numerous studies entailing the GHR signaling pathway have been conducted for various cancers. Diverse factors mediate the up- or down-regulation of GHR signaling through post-translational modifications. Of the numerous modifications, ubiquitination and deubiquitination are prominent events. Ubiquitination by E3 ligase attaches ubiquitins to target proteins and induces proteasomal degradation or starts the sequence of events that leads to endocytosis and lysosomal degradation. In this review, we discuss the role of first line effectors that act directly on the GHR at the cell surface including ADAM17, JAK2, SRC family member Lyn, Ubc13/CHIP, proteasome, βTrCP, CK2, STAT5b, and SOCS2. Activity of all, except JAK2, Lyn and STAT5b, counteract GHR signaling. Loss of their function increases the GH-induced signaling in favor of aging and certain chronic diseases, exempli

Looking for interaction: quantitative measurement of research utilization by Dutch local health officials

Background: In the Netherlands, local authorities are required by law to develop local health memoranda, based on epidemiological analyses. The purpose of this study was to assess the actual use of these epidemiological reports by municipal health officials and associated factors that affect this use.Method: Based on a conceptual framework, we designed a questionnaire in which we operationalized instrumental, conceptual, and symbolic use, the interaction between researchers and local health officials, and four clusters of barriers in this interaction process. We conducted an internet survey among 155 Dutch local health officials representing 35% of all Dutch municipalities. By means of multiple regression analyses, we gained insight into the related factors for each of the three types of research utilization.Results: The results show that local health officials use epidemiological research more often in a conceptual than an instrumental or symbolic way. This can be explained by the complexity of the local policy process which is often linked to policies in other areas, and the various policy actors involved. Conceptual use was statistically associated with a presentation given by the epidemiologist during the policy process, the presence of obstructions regarding the report's accessibility, and the local official's personal belief systems and interests originating from different professional values and responsibilities. Instrumental and symbolic use increased with the involvement of local officials in the research process.Conclusions: The results of this study provide a partial solution to understanding and influencing research utilization. The quantitative approach underpins earlier qualitative findings on this topic. The outcomes suggest that RPHS epidemiologists can use different strategies to improve research utilization. 'Blurring the boundaries', and the enhancement of interfaces between epidemiologists and local health officials, like direct interactions into each other's work processes, is expected to create better possibilities for optimizing research use

Knowledge in process? Exploring barriers between epidemiological research and local health policy development

The Redes de Trueque (RT) thrived during the economic crisis of 2001 – 2002 in Argentina and still stand out as one of the largest Complementary Currency System in the world. These local exchange networks reach a large scale during times of severe economic distress, but as large non-state initiatives, they pose a governance problem. Four types of governance systems were structured within the Argentine RT, of varying degrees of sustainability: a) loosely regulated market systems, b) hierarchies, c) associational regional networks, and d) local communities. Based on a four dimensional analytical framework, this paper discusses the rules of governance and sustainability of the governance systems in the RT. It found that some became more sustainable than others in terms of achieving combinations of scale and organisational modes

Growth Hormone Receptor Regulation in Cancer and Chronic Diseases: GHR Regulation in Cancer and Chronic Diseases

The GHR signaling pathway plays important roles in growth, metabolism, cell cycle control, immunity, homeostatic processes, and chemoresistance via both the JAK/STAT and the SRC pathways. Dysregulation of GHR signaling is associated with various diseases and chronic conditions such as acromegaly, cancer, aging, metabolic disease, fibroses, inflammation and autoimmunity. Numerous studies entailing the GHR signaling pathway have been conducted for various cancers. Diverse factors mediate the up- or down-regulation of GHR signaling through post-translational modifications. Of the numerous modifications, ubiquitination and deubiquitination are prominent events. Ubiquitination by E3 ligase attaches ubiquitins to target proteins and induces proteasomal degradation or starts the sequence of events that leads to endocytosis and lysosomal degradation. In this review, we discuss the role of first line effectors that act directly on the GHR at the cell surface including ADAM17, JAK2, SRC family member Lyn, Ubc13/CHIP, proteasome, βTrCP, CK2, STAT5b, and SOCS2. Activity of all, except JAK2, Lyn and STAT5b, counteract GHR signaling. Loss of their function increases the GH-induced signaling in favor of aging and certain chronic diseases, exemplified by increased lung cancer risk in case of a mutation in the SOCS2-GHR interaction site. Insight in their roles in GHR signaling can be applied for cancer and other therapeutic strategies

Jak2 inhibits GHR endocytosis.

<p>A. GHR-expressing Hek293 cells were transfected with empty vector (EV), Flag-Jak2, Flag-Jak2(Y119E) or Flag-Jak2(1-525). One coverslip was incubated for 15 min with Cy3-GH at 37°C, fixed and stained with anti-Flag (left panels). For EV- and Jak2-transfected cells, cells were also incubated with Cy3-GH and Alexa488-transferrin (Tf) for 15 min at 37°C (right panel). Bar, 5 µm. B. Hek293 GHR or GHR(UbE mutant)-expressing cells were transfected with Jak2 DNA or empty vector and, after 2 days, incubated for 2h on ice with 180 ng/ml ¹²⁵I-GH. To measure uptake kinetics unbound label was removed and the cells were incubated at 37°C for 10 min. C. Hek293 cells were transfected as in 1A and treated with GH for 15 min at 37°C as indicated. Total cell lysates (TCL, upper panel) and GHR immunoprecipitations (lower panel) were analysed on western blot (WB) using anti-Flag, or anti-phosphotyrosine (pY). 130, mature GHR; 110, precursor GHR. Data are representative of three independent experiments. D. Diagram showing the different binding motifs in the dimerised GHR; Jak2 binds Box1, TrCP binds UbE). The transmembrane domains are the dimerisation domains.</p

Decreased Jak2 activity in coated pit.

<p>A. GHR-expressing Hek293 cells were pretreated with MβCD for 1 h, incubated with Cy3-GH for 30 min at 37°C and the fluorescence was visualised with a confocal microscope. Bar, 5 µm. B. Cells were pretreated as in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0014676#pone-0014676-g004" target="_blank">Fig. 4A</a>, followed by incubation for 15 min with GH at 37°C as indicated. Total cell lysates (TCL, upper panel) and GHR immunoprecipitations (lower panel) were analysed on western blot with the indicated antibodies. 130, mature GHR; 110, precursor GHR. C. Cells were silenced either for clathrin or for GFP (as a negative control), followed by incubation for 15 min with GH at 37°C as indicated. Total cell lysates (TCL, upper panel) and GHR immunoprecipitations (IP, lower panel) were analysed on western blot with the indicated antibodies. 130, mature GHR; 110, precursor GHR. Data are representative for two independent experiments (B).</p

GH-induced and constitutive GHR endocytosis share the same characteristics.

<p>A. γ2A_GHR_Jak2 cells (left panel) and γ2A_GHR cells (right panel) were stimulated with 500 ng/ml GH for 90 min at 37°C as indicated. Total cell lysates (TCL) (lower panels) and GHR immunoprecipitates (upper panels) were analysed on western blot with the indicated antibodies. B. γ2A_GHR_Jak2 cells were transfected with control siRNA (lanes 1 and 2), clathrin heavy chain siRNA (CHC) (lanes 3 and 4) or TrCP siRNA (lane 5 and 6). Cells were stimulated with 500 ng/ml GH for 90 min at 37°C as indicated, after which total cell lysates (TCL) (3 lower panels) and GHR immunoprecipitates (upper panel) were analysed on western blot with the indicated antibodies. C. γ2A_Jak2 cells, stably transfected with wild-type GHR (WT), GHR mutated in its DSGxxS motif (DSG), GHR mutated in its UbE motif (UbE) and GHR with all cytosolic tyrosine mutated into phenylalanine residues (Y-less) were stimulated with 500 ng/ml GH at 37°C as indicated, after which total cell lysates (TCL) (2 lower panels) and GHR immunoprecipitates (upper panel) were analysed on western blot with the indicated antibodies. Cell lines used in C are stably transfected with the indicated GHR constructs, resulting in clonal variation of Jak2 levels. Data in A and B are representatives of three independent experiments. Data in C are representatives of two independent experiments. D. Quantification of the GH-induced GHR degradation (Y-less from experiment in C, and the data from the experiment in B), expressed as percentage of mature GHR.</p

mRNA levels of Jak2 and βTrCP2 in IM9, Hek293 and HepG2 cells.

<p>mRNA levels were measured using qRT-PCR. Ratios of Jak2/βTrCP2 are displayed of IM9, Hek293 and HepG2 cells as mean values of six independent experiments ± SD.</p

SCF(βTrCP) and Clathrin act downstream of Jak2.

<p>A. GHR-expressing Hek293 cells were either silenced for GFP (negative control), βTrCP or clathrin, and then treated for 15 min with GH at 37°C as indicated. GHR immunoprecipitations were analysed for ubiquitination. In the lower part of the figure the efficiency of silencing is visualised using anti-βTrCP or anti-Clathrin. B. Hek293 cells were transiently transfected with HA-tagged wild-type ubiquitin, ubiquitin K48R or ubiquitin K63R. Total cell lysates (TCL) and GHR immunoprecipitates were analysed on western blot using the indicated antibodies. EV, empty vector. C. Cells were transfected as in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0014676#pone-0014676-g001" target="_blank">Fig. 1A</a> and then treated for 15 min with GH at 37°C as indicated. GHR was immunoprecipitated from SDS-boiled lysates, and analysed for ubiquitination. 130, mature GHR, 110, precursor GHR. At the bottom of the figure the ratios of ubiquitinated versus 130kD-GHR are given. All data in this figure are representative of three independent experiments.</p